Submission of Pest Specimens for Identification
-Adapted from the Florida Forest Service Cooperative Forestry Assistance (CFA) Manual
Occasionally, foresters, land managers, or other individuals need the help of a specialist to identify a forest pest or its damage on host material. In some cases a field visit may be necessary, but in other cases, suitable specimen material may be shipped to our Forest Health Section office for identification. Please use the following guidelines to store and ship specimens in a way that will help prevent specimen damage and facilitate identification. This will allow us to make the best identification possible and optimize use of any stored specimen for future display or comparative purposes.
B. Specimen Collection and Storage
All insect specimens should be processed as soon as possible after collection to prevent loss of unique diagnostic characters, such as color, hairs, scales, appendages, and original body size.
Insects specimens may be sent in alcohol storage or dry-storage, depending on the insect species and life stage.
Alcohol Storage: Adult specimens of the following insect groups should be stored/shipped in alcohol. Many (but not all) of these insect groups are soft-bodied, small, and/or have few scales or hairs:
- Gnats (not mosquitoes)
- Tree Cattle
In addition, the immature stages (eggs, larvae, pupae, and some nymphs) of all insects should be stored/shipped in alcohol. A key point is that they are soft-bodied and unless placed in alcohol they will petrify, decay, and make identification virtually impossible.
Other arthropods (centipedes, millipedes, mites, spiders, ticks) should be stored/shipped in alcohol.
IMPORTANT HINTS FOR ALCOHOL STORAGE OF INSECTS:
- Grain alcohol (70-75%) is the best alcohol formulation to use for insect storage/shipment. A stronger concentration of alcohol, e.g., 95%, is not recommended because the insect specimens will harden after long-term storage. Isopropyl or rubbing alcohol, usually available in pharmacies and supermarkets, is suitable for use if grain alcohol is not available.
- All larvae (soft-bodied, worm-like, immature insects), e.g., caterpillars, maggots, grubs, should be immersed in boiling water for 1-5 minutes before final storage in alcohol. Boiling helps retain the color of the larvae; many will otherwise darken or fade in extended alcohol storage. Body size of the specimen will determine the length of immersion time: only 1 minute for small larvae (≤ 1 inch) and up to 5 minutes for large, robust larvae (≥ 2-3 inches). After allowing most water to drain off, place the specimen in alcohol. A pair of forceps or a small strainer will help minimize excessive handling of a specimen during this process.
- All storage/shipping containers, e.g., vials, bottles, jars, should be completely filled with alcohol. This will minimize the sloshing about of insect specimens which often have delicate appendages (antennae, wings, legs, hairs). Vials and bottles can be ordered through biological supply companies
- The best storage/shipping container is a glass vial topped with a tightly screwed cap or rubber/neoprene stopper. After inserting the specimen in the vial, completely fill it to the brim with alcohol to leave little or no air in the vial. Ideally, a label (written with pencil, which will not bleed in alcohol like ink) should be placed in the vial with the specimen. The label should at least specify the name of the collector, the date, and the county (or town) and state in which the specimen was collected. If using a screw-top cap, close the cap tightly and secure it with tape or other material that will help prevent leakage. If using a rubber stopper, place an insect pin or other slender pin/needle against the interior of the vial, and then press the rubber stopper deep into the vial while simultaneously withdrawing the pin. This procedure pulls surplus air out of the vial as a tiny stream of bubbles, helping to ensure an airtight fit.
- Do not place insect-damaged plant material and insect specimens in the same vial or bottle. The plant material will easily crush the insect specimens and/or decaying plant tissues may damage the specimens. However, insects firmly attached to plant material (e.g., scales, galls, cocoons) should be left intact on the plant and sent dry. In the case of firmly-attached scales or mealybugs, gently wrap the plant material in a dry paper towel and place it in a clear plastic Ziploc bag.
Procedures for packing a vial/bottle or loose plant material in a shipping container will be discussed in the “Shipment” section below.
Dry-Storage: Specimens of the following insect groups should be stored/shipped as dry-storage. Generally, these insects are hard-bodied, scaly, hairy, or large in size:
Remember, the immature stages of these groups are usually soft-bodied and require alcohol-storage (see above).
- Adult Flies and Mosquitoes
- Moths and Butterflies
- Lacewings, Antlions
- Dragonflies and Damselflies
- Grasshoppers, Crickets
IMPORTANT HINTS FOR DRY STORAGE OF INSECTS
- The less time spent handling a dry-storage specimen, the less likelihood of damaging it, i.e., breaking and losing important diagnostic characters: legs, wings, antennae, hairs, and scales.
- Killing jars, which are available at modest cost through biological supply catalogues, can be used to kill specimens destined for dry-storage. Common killing agents like ethyl acetate are safe if properly used by the collector. Some small insects can also be killed in a container placed in very cold freezer for a few days (but don't forget about them or they may become “freezer burned”).
- All dry-storage preparation should be done while the insect specimens are still fresh and pliable. Dried insects can be moistened by use of a relaxing chamber. This consists of a jar or bottle partially filled with sterilized, water-soaked sand. A small volume of a mild mold-inhibiting fungicide (i.e., Lysol aerosol) should be added to the sand. A specimen should not be allowed to directly touch the sand, rather place it in a shallow, open box. A dried specimen held for several hours in a tightly-closed relaxing chamber will allow limited rearrangement of an insect's appendages or body position.
- Before shipping for identification, dry-storage insect specimens should be placed in a small box or container and immobilized with tissue paper, cotton, or insect pins (very small pins, much like clothing pins. This small container should then be placed within a shipping container (see Shipment section below). If pinning the insect, push the pin vertically through the thorax (for most insects) and place the insect about 1 inch up on the pin, leaving enough pin sticking above the insect to grab easily. A piece of Styrofoam securely fit into the bottom of the small (interior) container makes a good base for the pin. See the Resources section below for more help in this area.
- If you are keeping dry storage insect for an extended period of time, moth balls or flakes (commonly naphthalene or paradichlorobenzene) should be used to preserve them. Neither material should be allowed to directly contact specimens. The best large storage containers will have a tight seal to (1) prevent entry of other insects (silverfish, carpet beetles) that feed on stored specimens and (2) allow permeation and retention of the evaporating preservatives. If you are shipping a dry-storage specimen by mail for identification, you do not need to include these preservative materials.
Effective and accurate pathological diagnoses are dependent upon the acquisition of appropriate sample/specimen materials which are carefully handled so as to retain, as much as possible, their key diagnostic features. The most common disease specimens collected, stored and shipped are portions of plants or trees exhibiting various symptoms of pathogenic infection, e.g., galls, cankers, leaf spots, sap exudation or resinosis, tissue necrosis, decay. Frequently, the sporocarps (fruiting bodies such as conks and mushrooms) of suspect pathogenic fungi may also be observed in association with diseased trees or tree parts, and thus are collected, packaged and forwarded to diagnostic laboratories. The following comments are presented as guidelines for aiding in the proper collection of tree disease specimens.
Collection of Samples:
|Type of Problem
|I. LARGE TREES
A. Localized gall, canker, leafspot, etc.)
a) 3-4 branch-portions or ca. 10-12 leaves exhibiting symptom(s) of concern. If possible, obtain samples exhibiting symptoms in various stages of development.
b) Associated fungus sporocarps, e.g., conks, mushrooms
B. General (Chlorosis, Wilt, Crown thinning, Dieback, etc.)
a) Soil: from upper 6” of soil in root zone; ca. one pint (total) from 5-10 locations within dripline of tree’s crown, include small feeder roots if possible.
b) Foliage: ca. 1-2 dozen leaves
c) Branch: 4-6 branch portions between ½ and 2” in inch diameter and 6-10” inch length (especially branches with evidence of canker infections, vascular discoloration, etc.)
d) Roots: 3-4 segments between ½ and 2” inch in diameter and 6-10” inches in length only if they exhibit obvious internal discoloration, resinosis, or decay; not always obtainable in urban settings
e) Associated fungus sporocarps, etc.
a) 3-6 “diseased” seedlings (entire seedling) together with ca. two cups of soil from root zones; prefer seedlings packaged individually
b) 3-6 “healthy” seedlings with soil for comparison. Must be packaged separately from diseased specimens.
IMPORTANT HINTS FOR DISEASE SPECIMEN COLLECTION
- Pathogens are living organisms.
- Pathogens are usually located in and cultured from the transition zone between healthy and “diseased (necrotic, stained, discolored, etc.) tissues.
- Do not submit only long dead, bone dry material to a pathology laboratory for cultural diagnosis. Some of this material may be useful, but material in the process of dying or developing symptoms is essential.
- Pathogens are killed by excessive heat and drying, thus preventing successful isolation/detection and diagnosis in the laboratory. Collected specimens should be kept cool and moist (not wet).
Packaging & Storage of Disease Specimens:
Packaging of disease samples/specimens is a relatively straight forward task. Most specimens can simply be placed in a sufficiently large clear plastic bag, labeled appropriately (either by writing on the bag with a water insoluble, felt tip marker or by attaching some type of label, tape, etc.) and sealed. Zip lock-type bags are very convenient, although regular plastic bags are equally suitable. Specimens subject to excessive dehydration or desiccation (leaves, small twigs or small roots, etc.) should be packed with a moist (not soaked) paper towel to facilitate moisture retention. Large root and branch samples, as well as soil samples and small roots packed with soil, do not require and generally should not be packaged with additional moisture.
Short-term storage of disease specimens is best accomplished by refrigeration. Ice chests for field storage and transport of specimens are most helpful. If long term storage of specimens is desired, a variety of methods are available, depending upon the type of specimen(s), available space, objectives of the collector, available monies, etc. Shoe boxes containing 1-2 moth balls work well for certain types of dried materials. Sometimes it may prove useful to package dried specimens in plastic bags with 1-2 moth balls before storing in larger, “collective” boxes Pressed and dried leaves are often stored in Riker Mounts (glass-faced, cotton-backed boxes ideal for display, but expensive). Fleshy fungi (mushrooms, toadstools, etc.) require careful and rapid drying, during which they often lose their color and shrivel considerably prior to storage.)
C. Shipment of Specimens
According to USPS publication #52, Hazardous, Restricted, and Perishable Mail, the Hazard Class of Isopropyl Rubbing Alcohol (70%) is 3. A flashpoint of 53* F qualifies rubbing alcohol as a “Flammable Liquid”. Flammable liquids are prohibited from domestic air transportation, and all international shipping. However, it may be shipped domestically by ground, but only in limited quantities according to specific packaging instructions and labeling. Shipping instructions are as follows:
- Domestic ground transportation only!
- Limited quantity =< 30ml (1 oz.)
- A metal primary receptacle must not exceed 1 quart. (This is the container size. Alcohol content is limited by above.)
- A nonmetal primary receptacle must not exceed one pint.
- The primary receptacle must have a screw cap (with a minimum of one-and-one-half turns), soldering clips, or other means of secure closure (friction caps are not acceptable).
- Only one primary receptacle is permitted per mailpiece.
- Enough cushioning material must surround primary receptacle to prevent breakage and absorb leakage.
- The cushioning material and primary receptacle must be packed in a securely sealed secondary packaging.
- A strong outer packaging that is capable of firmly and securely holding the primary receptacle, cushioning material, and secondary packaging is required.
- Each mailpiece must be clearly marked on the address side with “Surface Mail Only”, “Consumer Commodity ORM-D”, and the flashpoint of the flammable liquid (53* F).
- A complete return address and delivery address must be used.
Specimens shipped dry or in alcohol should be placed in a large container (e.g., cardboard box, mailing tube) with firm sides that will resist crushing from improper handling. Dry-storage insect specimen containers, plant material, vials, and bottles need to be immobilized within the shipping container. Common packaging materials are loosely wadded newspaper, paper toweling, tissue paper, and small pieces of Styrofoam. Before sealing the shipping container, make sure that a completed Specimen Report Form (see below) is included for each type of specimen. Avoid sending specimens on a Thursday or Friday, if possible, to prevent specimens sitting at room temperatures over weekends.
Ship the container to the attention of the appropriate pest specialist (Forest Entomologist, Forest Pathologist, or Invasive Plant Biologist) at the following address:
Forest Health Section
Florida Forest Service
1911 SW 34th St.
Gainesville, FL 32608
D. Specimen Report Form
When submitting specimens for identification or diagnosis, please print and fill out a Division of Plant Industry Specimen Report Form for each specimen submitted.
E. Resources for Collecting and Storing Specimens
- A fantastic, comprehensive online reference for insect collecting, preserving, mounting, and shipping is “Collecting and Preserving Insects and Mites” by the USDA Systematic Entomology Library.
- A good shelf reference for insect collection, preservation, storage (as well as identification) is A Field Guide to the Insects of America North of Mexico by D.J. Borror and R.E. White (part of the Peterson Field Guide Series).